Project Summary. The world is heterogeneous with respect to danger and safety. While it is adaptive to be afraid when confronted with certain danger, an equivalent level of fear in safety or even uncertainty is maladaptive and detrimental to health. Precise control of fear is disrupted in anxiety disorders, such as post-traumatic stress disorder, with those affected displaying exaggerated fear to safety cues. The goal of this project is to uncover how RRF neural activity permits precise control of fear. To do this, we have designed a novel and challenging fear discrimination procedure. In fear discrimination, three auditory cues are associated with different probabilities of foot shock: danger (p=1.00), uncertainty (p=0.25), and safety (p=0.00). Cue presentation is either random or blocked. Rats normally show precise control of fear as indicated by high fear to the danger cue, intermediate fear to the uncertainty cue, and low fear to the safety cue. In Specific Aim 1, we test the hypothesis that phasic RRF activity reflects an estimate of safety while tonic RRF activity reflects an estimate of danger. We will implant drivable microelectrode bundles in the RRF of adult male and female rats, and RRF activity will be recorded during fear discrimination. Analysis of phasic activity will focus on the cue presentation period. We predict that a significant population of RRF neurons will phasically increase firing in accordance with an estimate of safety: large phasic increases in activity to the safety cue, intermediate increases to the uncertainty cue, and low/no increases to the danger cue. Analysis of tonic activity will focus on the time period between trials. We predict that a significant population of RRF neurons will alter tonic firing levels in accordance with the current block: highest tonic RRF activity will be observed in the danger block, intermediate tonic activity in the uncertainty block, and lowest tonic activity in the safety block. In Specific Aim 2, we will use optogenetic inhibition and excitation to provide causal roles for phasic RRF activity underlying an estimate of safety and tonic RRF activity underlying an estimate of danger. Inhibition will be achieved by transfecting RRF neurons with viral constructs containing halorhodopsin, implanting optical ferrules above, and illuminating the RFF with green light. Excitation will employ the same general strategy, but will use channelrhodopsin in combination with blue-light illumination. Effects of RRF inhibition or excitation in the cue period will be compared to periods outside of cue presentation. We predict that phasic inhibition or tonic excitation of RRF activity will increase fear to all cues. By contrast, phasic excitation or tonic inhibition of RRF activity will decrease fear to all cues. Cumulatively, Aims 1 and 2 will demonstrate that RRF activity on multiple time scales is essential to the appropriate display of fear. The results of this proposal will uncover the RRF as a key brain region permitting the precise control of fear, as well as a locus of dysfunction and a compelling pharmacotherapeutic target for psychiatric disorders in which control of fear has been lost.